Ординатура / Офтальмология / Английские материалы / Basic Sciences in Ophthalmology_Velayutham_2009

abundant there. Most GAGs occur linked to core proteins, the entire assembly is known as proteoglycan. Proteoglycans and GAGs are often associated with collagen in extracellular matrices. The common GAGs found in ocular tissues are:

Hyaluronic Acid

Composed of Glucuronic acid and N- acetyl glucosamine linked together by ß 1-3 glycosidic linkage within the unit and β 1-4 glycosidic linkage in between units.

Chondroitin Sulfate

Glucuronic acid and N- acetylgalactosamine sulfate in ß 1-3 glycosidic linkage.

Keratan Sulfate

Galactose and N–acetyl glucosamine sulfate in 1-4 glycosidic linkage.

Dermatan Sulfate

Iduronic acid and N acetyl galactosamine sulfate in β 1-3 glycosidic linkage. Sulfation of the right hand unit adds considerable acidity and charge density to the entire unit. The negative charge density is an important characterstic of

GAGs that attract counterions ( principally sodium ) and water.

The major function of structural carbohydrates is to:

•Increase the stability of protein (hydrophilic).

•Stabilise protein conformation.

•Aid in proper orientation of protein in membrane.

•Act as recognition marker for cell sorting prior to protein transport.

•Acts as immunological identifiers for immune reaction.

Proteoglycans

The core proteins to which GAG is bound to form the proteoglycans are:- Keratocan, mimican, lumican—bound to keratan sulfate and decorin – bound to dermatan sulfate.

One to 3 GAGs are bound to core protein through oligosaccharide chain to form the proteoglaycan. The link is N glycosidic bond between N-acetyl glucosamine of keratan sulfate and asparagine in core protein (Fig. 12.8).

Fig. 12.8: Proteoglycan

LIPIDS

Are the structural basis of cell membranes. Membrane is composed of phospholipids, glycolipids, cholestrol etc.

Phospholipids

Are the most important lipids for the formation of maintainance of cell membrane. In this, glycerol is used as the frame on which 2 fatty acid esters are attached to the 1st and 2nd carbon and a phosphate ester to the 3rd carbon to which any of the 4 polar groups - ethanolamine, choline, serine or inoistol is attached, e.g. Phophotidyl choline (Fig. 12.9).

In an aqueous medium, lipids arrange themselves in such a way that the polar (hydrophilic) groups face the aqueous phase, whereas the hydrophobic groups face each other to form a miscelle (Fig. 12.10). But owing to the bulky nature of 2 chains of fatty acids, membrane lipids do not readily from the miscelles,but group together to form a lipid bilayer (Fig. 12.11).

Phospholipid bilayers have the fatty acid composition designed by the cell as per its functional need. For, e.g: RBC must have somewhat a rigid membrane to assume its biconcave disc shape. So, it tends to have a shorter chain with unsaturated fatty acids and a lower percentage of long chain, highly unsaturated fatty acids. But rod outer segment discs require a high degree of membrane fluidity to carry out the process of visual transduction. So, higher percentage of long chain unsaturated fatty acid is required. Cervonic acid – (a 26 carbon fatty acid with 6 double bonds) is one such fatty acid.

Cholesterol is also a component of membranes. It is hydrophobic predominantly except for the hydroxyl group on 3rd carbon. It gives rigidity to the membrane so, the rod outer segment has lower percentage of cholesterol ie only 8%.

Fig. 12.9

Fig. 12.10: Miscelle

Fig. 12.11: Lipid bilayer

Glycolipids

Glycolipids are another important membrane components found in ocular tissues. They contain carbohydrates like galactose, N- acetyl galactose and N- acetyl neuraminic acid (sialic acid). The basic structure that binds the glycolipids is not glycerol, but a long chain amino alcohol known as sphingosene.

Sphingosene + fatty acid → ceramide.

Ceramide + phosphatidylcholine → sphingomyelin.

If phosphocholine is replaced by carbohydrate, it is glycolipid or glycosphingolipid. They are cerebrosides and gangliosides. Apart from these above mentioned lipids, there are some derived lipids called eicosanoids. They are cyclic lipids derived from eicosonic acid such as arachidonic acid. They include prostaglandins, thromboxanes and leukotrienes which are short acting local hormones. They are formed from membrane phospholipids by the hydrolytic action of the enzyme phospholipase A2.

Synthesis of Eicosanic Acids

Prostaglandins

Flowchart 12.1

Intiation – production of free radicals or reactive oxygen species as correctly named viz R· and ROO·.

R° is called the carbon centered radical. ROO· is the lipid peroxide radical. Interaction of PUFA with free radicals generated by other means

(such as univalent reduction of oxygen in some enzymatic reactions and by metal ions) leads to the formation of R· and ROO·. Free radicals are molecules or atoms that have an unpaired electron which makes them highly reactive.

H2O

E.g. superoxide (O2·) hydroxyl radical (OH·) hydrogen peroxide (H2O2). R· and ROO· in turn are degraded to injurious products.

ROOH + metal ——→ ROO· + H+ RH (Fatty acid) + OH· → R· + H2O.

Propagation Phase

R° rapidly react with molecular oxygen to form peroxyl radical (ROO°) which can attack another polyunsaturated lipid molecule and the reactions continue to produce peroxyl radical from PUFA of membrane destroying the integrity of the membrane.

R· + O2 ——→ ROO·

ROO· + RH (Fatty acid) → ROOH + R·

Termination Phase

When 2 peroxyl radicals react with each other inactive products form, thus terminating the damaging chain reaction.

ROO·+ ROO· → RO – OR + O2 R· + R· —→ R–R

ROO· + R· → RO – OR

The oxidation of membrane phospholipids is thought to increase the membrane permeability of cell and inhibit membrane ion pumps. This loss in barrier function leads to edema, disturbances in electrolyte balance, elevation of intracellular calcium, all resulting in malfunctioning of cell.

The damage produced by reactive oxygen species may be prevented by antioxidants which are of 2 types –preventive and chain breaking.

Preventive antioxidants inhibit the initial production of free radicals, they are catalase, glutathione perioxidase trdiamine diethyl penta acetate (DTPA) and ethylene diamine tetra acetate (EDTA). Once the peroxyl radicals are generated, the chain breaking antioxidants inhibit the propagative phase. They include superoxide dismutase, Vit E and uric acid. Vit E (alpha tocoferol T- OH) would intercept the peroxyl free radical and inactivate it before a PUFA can be attacked.

glucose transporters (GLUT). GLUT allows the diffusion of glucose into the cells down their concentration gradient. GLUT are proteins found in cell membranes with 12 transmembrane domains. They are of 5 types — GLUT 1- GLUT 5. They are all facilitative bidirectional transporters. SGLUT1 alone is sodium dependant unidirectional transporter, which is an active mechanism against concentration gradient. Some are insulin dependant or regulated by insulin. E.g: GLUT 4 others are not regulated by insulin. The GLUT in ocular tissue is mainly GLUT1 andGLUT 3.

Glucose on entering the cells through GLUT undergoes many types of reactions for varied uses in the cells.

The initial reaction is the conversion of glucose to glucose 6 phosphate, utilizing energy in the form of hydrolysis of a high energy compound, ATP. This is to prevent the escape of glucose from the cell. The negative charge on the phosphate group will not allow the glucose to pass through the hydrophobic interior of the plasma membrane of the cell.

The enzyme, hexokinase catalyzing this reaction has high affinity for glucose and other enzyme glucokinase, catalyzing the same reaction has low affinity for glucose. So only when the glucose concentration is high, glucokinase will act.

Glucose 6 phosphate is the metabolic junction for many pathways-viz Glycolysis – Aerobic for production of enrgy in the form of ATP

– Anaerobic

Pentose phosphate pathway – for production of NADPH and pentoses on Glycogenesis for storage of energy.

Glycolysis

Glucose 6 phosphate is prepared for fractionation into 3 carbon units,split apart then further rearranged to generate 4 molecules of ATP.

Phosphoglucoisomerase

Glucose 6 phosphate ———————————— fructose 6 Phosphate

The next reaction energetically primes the molecule for splitting

Upto this reaction, 2 molecules ofATP have been consumed and 4 molecules of ATP have been formed with the net gain of 2 ATP per glucose molecules. Pyruvate is at the junction point between aerobic and anaerobic metabolism. In anaerobic metabolism, there is only a single reaction that forms lactate from pyruvate and with that the pathway is terminated. The NADH formed in the earlier reaction is utilized here, instead of entering, etc.

This is a quick and relatively uncomplicated means for cells to obtain ATP in the absence of oxygen, though the yield is small ( 2 ATP/ glucose molecule). If the cell obtains its glucose from the breakdown of stored glycogen, it usually realizes a netgain of 3 ATPs anaerobically because no ATP is required for formation of glucose 6 phosphate from glycogen.

The cell obtain high energy supply in a short period. So, a relatively significant percentage of glucose is utilized via this pathway in ocular tissues. Moreover, sufficient amount of NAD must be regenerated from NADH for use in the earlier reaction, i.e. formation of 1,3 bisphosphoglycerate from glyceraldehyde 3 phosphate for that purpose also, this anaerobic glycolytic pathway is necessary.

Aerobic Glycolysis

Pyruvate, if not converted to lactate, will diffuse into cellular mitochondria, to begin the aerobic phases of ATP production (the mitochondria has a double membrane, the inner membrane has a large surface area with infoldings known as ‘crista’ and it is impermeable to most of the molecules and ions without a transporter. It contains a number of insoluble electron transfer proteins and an enzyme, ATP synthase. The inner most compartment called “matrix” contains many soluble enzymes not found in cytoplasm).

Pyruvate is converted to acetyl CoA in the matrix and the complete oxidation occurs in matrix. The reducing equivalents that are produced during oxidation, enter the respiratory chain or electron transport chain in the inner membrane to produce ATP and CO2 which is used for formation of oxaloacetate from pyruvate.

TCA cycle (Fig. 12.12)

Respiratory chain or Electron transport (ETC) is the site of ATP production during oxidation of the reducing equivalents.

Fig. 12.12

ETC

NADH + H+ ——————NAD + H2O +3 ATP

O2

ETC

FADH2 ——————— FAD + H2O + ATP O2

Substrate level phosphorylation (ATP from ADP) during conversion of succinyl CoA to succinate. Totally 12 ATPs are produced per citric acid cycle (TCA cycle). Thus, on complete oxidation of one molecule of glucose 36/38 ATPs are produced aerobically.